1. Field of the Invention
The present invention relates to a fluorescent microarray analyzer, particularly to a fluorescent microarray analyzer for detecting fluorescent signals emitting from a biochip by means of spectrum analysis.
2. The Prior Arts
As “working draft” of the human sequence unraveled by the Human Genome Project published in Nature (15 Feb., 2001), simultaneously with a companion publication of the human sequence generated by Celera Genomics Corporation (Science, 16 Feb., 2001), understanding the physical functions and the mechanisms of human genes becomes the next important goal in the field. To accelerate the progress of the related research, high-throughput tools for efficient analysis are available. Biochip, results of mass of samples expressed on surface of a small solid carrier, is such a useful analytic tool. Biochip can be employed in gene expression, drug selection and disease diagnosis in both basic research and clinical application fields.
Three kinds of biochips are known, namely DNA chip, lab-on-a-chip and protein chip. Since the protein chip and the lab-on-a-chip are difficult to operate, the DNA chip is in common use now. The detection of the DNA chip is shown in FIG. 1. Known DNA fragments serving as DNA probes (2) are immobilized onto a surface of a glass slide or a silicon chip and form a DNA chip (1). Generally, the DNA probes (2) are arranged in array, which is called DNA microarray. On the other hand, unknown DNA fragments (3), the target DNA, are labeled with fluorescent dyes. The DNA chip (1) is then hybridized with the target DNA (3). After washing, only DNA fragments hybridized with DNA probes are left on the DNA chip (1). By scanning with a biochip reader, the fluorescence from the fluorophores on the slide is detected and the obtained hybridization result is analyzed.
FIG. 2 is a schematic view showing a conventional biochip reader (4). Beams of light emitted from a laser source (40) pass through a focusing lens (41), reflected by a beam splitter (42), and then further passing through another focusing lens (43) to irradiate a surface of the biochip (44). The fluorescent dyes on the biochip (44) are excited by the beams of light and in turn emits fluorescence (45). The fluorescence (45) passes through the focusing lens (43), the beam splitter (42), and the focusing lens (46). The fluorescence (45) further passes through a filter (47) and with which the beams of light from the light source are filtered out. The fluorescence is then detected by a photomultiplier tube (PMT) (48), which converts the optic signals into electrical signals, which are fed to a computer (49) and processed to form image data. In the conventional biochip reader, to obtain the final result requires scanning all samples on the biochip, converting optic signals into electrical signals, and forming the electrical signal image data for analysis. The conventional biochip is disadvantageous, since errors occur in the formation of the image data by processing the electrical signal and it takes much time to obtain the image.
Cyanine 3 (Cy3) and cyanine 5 (Cy5) are two fluorophores in common use. Cy3, with peak absorption at 550 nm, is generally excited with a laser beam of 532 nm and emitting fluorescence which has central wavelength at 570 nm. Cy5, with peak absorption at 649 nm, is generally excited with laser beams of 632.8 nm or 635 nm and emitting fluorescence which has central wavelength at 678 nm. Filters are generally used to eliminate the interference caused by the original beams of light from the light source and the scattering light from the slide. When filtering out the interfering light from the light source, some signals of the resulting fluorescence may be filtered out at the same time due to the crosstalk between the fluorescence and the interfering light. In addition, problems of exchanging filters happens when two or more fluorophores are used on the same biochip.